LAUSR

Abstract Germ cell transplantation (GCT) is a promising reproductive technology in animals, including teleost fish. Here, we established a GCT procedure in Japanese flounder (Paralichthys olivaceus). Two-year-old male Japanese flounder were treated with temperature (28 °C) and busulfan (two doses, 18 mg/kg) to deplete endogenous germ cells and were then used as recipients. The discontinuous Percoll gradient purified oogonia were used as donor germ cells and transplanted into the recipients' testes through the genital pore. Histological and fluorescent observations showed the colonization, proliferation, and differentiation of transplanted oogonia in the testes of recipient fish. Ten months after GCT, 16 of 17 recipients produced sperm, and among them, two recipients produced donor-derived sperm, which was confirmed by microsatellite genotyping. The sperm of these two recipients were used to fertilize the eggs of wild-type females, and the donor-derived germline transmission rates were 45.5% and 43.0%, respectively, based on the microsatellite paternity analysis of the hatched larvae. Our established protocol can be applied to accelerate the reproduction cycle and be used for generating valuable surrogate breeding material and conserving the germplasm resources of Japanese flounder.